The International Agency for Research on Cancer (IARC) recognized the Epstein-Barr virus (EBV), hepatitis B virus (HBV), hepatis C virus (HCV), Kaposi’s sarcoma herpes virus (or human gammaherpesvirus 8), human immunodeficiency virus 1 (HIV-1), several human papillomavirus (HPV) types, and human-T lymphotropic retrovirus-1 (HTLV-1) as biological agents involved in human carcinogenesis. Criteria used to prove the involvement of these viruses in the tumorigenesis process was based on analyzing exposure data, studies on cancer in humans and in animal models, and identification of relevant data providing mechanistic insight. Exposure data refer to general information about the agent, analysis and detection methods regarding sensitivity and specificity, occurrence, and exposure. Research of cancer in humans analyzed the type of studies (cohort studies, case-control studies), meta-analyses and pooled analyses, temporal effect, the use of biomarkers in epidemiological studies, and criteria of causality. Model animal studies investigated the qualitative and quantitative aspects, and mechanistic insight referred to toxicokinetic data, mechanisms of carcinogenesis which identified functional changes at the cellular level and alterations at the molecular level [1].

The estimated age-standardized incidence rates in 2020 for skin melanoma and non-melanoma skin cancer, both genders, all ages, WHO Europe, mention Switzerland, Ireland, and The Netherlands in the first three places, with age-standardized rates (ASR) 71.1, 63.3, and 61.5, respectively, with Romania having an ASR of 12.0 [2]. This data collection regarding the incidence of cancer is powered by population-based cancer registries which are available in developed countries; this led to the idea that in countries without population-based cancer registries, the incidence of cancers is underreported. Skin cancer is known to have several risk factors, such as sun exposure, BRAF mutation in melanoma patients, and some molecular factors ^[3][4][3,4]. The higher incidence of NMSC (nonmelanoma skin cancer) in immunocompromised patients points to a possible viral origin [5].

The Polyomaviridae family includes numerous small, icosahedral, non-enveloped viruses, which have a double-stranded DNA genome that is approximately 5000 base pairs in length, and it is packed together with histones uptake from the host cells. These viruses have a wide range of hosts, including mammals, birds, and fish [6]. The International Committee on Taxonomy of Viruses (ICTV) currently recognizes eight different genera of polyomaviruses (Alpha-, Beta-, Gamma, Delta-, Epsilon-, Zeta-, Eta- and Thetapolyomavirus), comprising a total of 117 species [7]. The genetic diversity of these viruses is also very great, and a characteristic co-speciation with their hosts has been observed, which is a result of genetic recombination, as it has been observed for papillomaviruses [8]. Many of the viruses in this family are associated with an oncogenic capacity in animal hosts, which has been observed since the discovery of murine polyomavirus in the 1950s. The Polyomaviridae family was given this name because of the numerous types of tumors they can induce (polyoma) [9].

Of the human polyomaviruses, MCPyV was the first one for which evidence of carcinogenic potential has been observed, in a rare and aggressive form of skin cancer named Merkel cell carcinoma (MCC). MCPyV was first discovered at the Pittsburgh Cancer Institute in 2008, using digital transcriptome subtraction assays. The researcheuthors detected that the viral DNA integrated within the tumoral cells’ genome in a clonal pattern, in 6/8 MCPyV-positive MCCs, suggesting that MCPyV infection and integration preceded clonal expansion of the tumoral cells. MCPyV was then first considered to have a contributing factor in the pathogenesis of MCC [10]. Four years later, scientists from 11 countries met at IARC, to evaluate the carcinogenicity of MCPyV, and their research has been published in a monograph and in a Lancet paper. By analyzing all the research studies published since its discovery, the researcheuthors concluded that there is powerful mechanistic evidence that MCPyV can directly contribute to the development of a large proportion of MCCs. Using PCR, many independent laboratories have detected MCPyV DNA in about three quarters of more than 1000 MCC cases ^[11][12][11,12].

In 2017–2018, a multidisciplinary team from important research centers of many continents (e.g., German Cancer Research Centre, DKFZ, Heidelberg, Germany, Department of Melanoma Medical Oncology, Division of Cancer Medicine, MD Anderson Cancer Centre, Houston, TX, USA, Peter McCallum Cancer Centre, Melbourne, Australia, to name just a few) published three reviewsearch regarding MCPyV in relation to skin cancer ^[13][14][15][13,14,15].

Becker JC et al., classified MCCs in MCPyV positive and negative and mentioned that in the countries with low UV exposure, MCPyV is present in most of the skin cancers, in stark contrast with countries with high UV exposure where the virus is absent in MCCs. It is interesting that both MCC types have similar phenotypes, and several tissue markers were detected in skin cancer that may be positive or not for MCPyV, including apoptosis regulator B-cell lymphoma-2 (BCL2), cytokeratin 20, neural cell adhesion molecule 1, CD99, CD99 antigen, epithelial cell adhesion molecule, huntingtin-interacting protein 1, neuron-specific enolase, and neurogenic locus notch homologue protein 1. The MCPyV-specific MCC viral markers are large T antigen and small T antigen. The researcheuthors mentioned that, in the case of viral-positive MCC cases, the genetic aberrations observed are from perturbations of signaling pathways by antigens and genome integration; meanwhile, in the case of UV exposure, other alterations were detected, such as deletions, translocations, and point mutations [13].

In the second resviearchw [14], members of the EU IMMOMEC (European Union Immune Modulating strategies for treatment of Merkel Cell Carcinoma) presented the actual available therapy that is efficient for this type of skin cancer: the immune checkpoint-inhibiting antibodies pembrolizumab and avelumab [specifically, the programmed death protein 1 (PD-1) and programmed death-ligand 1 (PD-L1) blocking antibodies]. This new therapy seems to be efficient in more than half of the treated MCC patients. Still, a targeted therapy is still necessary, as many MCC patients are immunosuppressed and their response to immune checkpoint inhibition is not possible [15].

In 2018, the International Workshop on Merkel Cell Carcinoma Research (IWMCC) working group underlined some open research questions regarding this primary cutaneous neuroendocrine carcinoma, MCC: the multidisciplinary research team (e.g., virology, pathology, oncology, dermatology) raised awareness regarding future targeted therapy in both MCPyV positive and MCPyV negative cases of MCC, and about the optimal detection assay for this virus [15].

The Papillomaviridae family is comprised of small, icosahedral, non-enveloped viruses with a double-stranded DNA genome, and are also characterized by a great genetic diversity and wide range of hosts, including mammals, birds, reptiles, and fish. They also have a known oncogenic potential in humans, most importantly in the development of cervical cancer, but also vulvar, vaginal, penile, and oropharyngeal cancers. The human papillomaviruses which are associated with those cancers are also called mucosal, high-risk, or alpha HPV types [16].

The first classification of cutaneous papillomaviruses was performed by de Villiers EM et al., in 2004 [17]. In 2012, the IARC monograph reported that, up to that moment, there was no HPV type which could be considered to cause skin cancer, due to the lack of consistency of the published data. In 2012, it was considered that the role of HPV types in skin cancers could be complex, possibly associated with other co-factors, such as UV exposure [1]. In 2013, over 170 human papillomavirus types were reported to be associated with different clinical manifestations in humans, with the skin being the main site, followed by mucosa (vagina, mouth) and gut [18].

In a similar manner to MCPyV, for cutaneous HPV types, important reviesearchws were recently published, with a different view regarding the involvement of these viruses in skin cancer.

Venuti A et al., analyzed the “cross-talk” between cutaneous HPV types and the immune system, in a journal of the Royal Society; they mentioned the “hit-and-run” hypothesis, having the ability to initiate the first steps of UV-driven skin carcinogenesis, a different mechanism of carcinogenesis, in comparison with that of mucosal HPV types responsible for cervical cancer. The researcheuthors underlined the necessity of understanding the cross-talk with host cell-autonomous and extrinsic immunity for it to be possible to identify novel therapies against beta HPV, besides their sensitivity to interferon regulatory factors [19].

Gheit T., 2019, an IARC researcher with impressive experience in HPV testing and analyzing, presented the main features and functions of the early and late gene products from alpha and beta HPV types. Interestingly, for E6 and E7 genes known as oncogenic in cervical cancer, different functions are underlined: both genes are not required for the maintenance of the cancer phenotype. E6 interacts with the Notch pathway and promotes the transformation process of the infected keratinocytes, and inhibits the differentiation of HPV8-expressing keratinocytes by targeting the PDZ domain-containing protein syntenin 2. E7 from HPV38 shows the ability to counteract p53-mediated apoptosis by inducing an accumulation of the p73 isoform, 1Np73 [16]. The researchersuthor was in support of the hypothetical carcinogenesis mechanism of the research^[19]previous review [19], mentioning that E6 and E7 expression appear to be required only at the initial step of skin carcinogenesis by exacerbating the deleterious effects of UV radiation [16].

Rollison DE et al., 2019, mentioned that since the first meeting group at IARC regarding beta HPV types, 50 types of cutaneous HPV have been identified from a total of 200 HPV types. The researcheuthors underlined the importance of UV as a co-factor in skin carcinogenesis, in the case of constant stress, and it is considered that cutaneous HPV types facilitate DNA damage accumulation induced by UV radiation. This review used high-risk HPV (HR-HPV) types as a comparison, and the researcheauthors are confident that, if for cervical cancer developing three vaccines (bi, tetra, and nonavalent) was possible, it will be feasible to create a vaccine against beta HPV types [20].

Given these recent data regarding skin cancer and the association with two potentially oncogenic viruses, rwesearchers aim in this review to present updates regarding detection methods, carcinogenetic mechanisms, and the availability of therapeutic vaccination for MCPyV and cutaneous HPV types.

ItWe is analyzed the articles published in the last 5 years using the following keywords:”skin cancer Merkel cell polyomavirus detection assay”. From all 34 studies suitable for theour research, only 13 studies were selected. Since 2007, the detection methods for Merkel cell polyomavirus have evolved from serological diagnosis MCV-oncoprotein antibody detection [21], to testing by simultaneous complementary molecular techniques (classical and qPCR) ^{[22][23][24][25][26][27]}[22,23,24,25,26,27], or by double analyzing MCC with molecular and immunohistochemical analyses [28]. This double testing underlines the necessity of confirmatory diagnosis by two different or complementary assays. An interesting study performed in France analyzed the circulating tumoral cells (CTCs) in blood samples for patients with MCC, and the researcheuthors remarked on the tumor heterogeneity [29]. A very comprehensive study was the one realized at Bethesda, MD USA, in 2020, in which the researcheuthors used deep sequencing with OncoPanel, a clinically implemented, next-generation sequencing assay targeting over 400 cancer-associated genes; they observed the value of high-confidence virus detection for identifying molecular mechanisms of UV and viral oncogenesis in MCC [30]. MCV DNA was detected in formalin-fixed and paraffin-embedded (FPPE) MCC samples, with different prevalence, from 10% up to 90%. The most common primers used were targeting the sT gene, VP1 and NCCR regions of the genome, and large T-antigen (LTAg) gene; the sequence of used primes are presented by the researcheuthors ^{[21][22][23][24][25][26][27][28][29][30][31][32][33]}[21,22,23,24,25,26,27,28,29,30,31,32,33].

Oncogenic transformation by MCPyV is hypothesized to require two events: the integration of the viral genome into the host genome, and the truncation of large Tantigen to render the viral genome incapable of replication. In the viral positive MCCs, the small T antigen has an important role in carcinogenesis: it is known to transform rat-1 fibroblasts in culture. Research carried out on transgenic mouse models has shown that the expression of small T antigen was transformative in various organ systems, including in the epidermis [15].

The carcinogenetic mechanisms identified from 2017 to 2022 were determined in different modalities, beginning with evaluating the MCPyV cultivation on cell lines, to assessing the expression of specific genes. The outcomes of the analyzed studies were correlated with possible future targeted therapies, including for metastatic MCC, and even with future vaccinations ^{[34][35][36][37][38][39][40]}[34,35,36,37,38,39,40].

Skin biopsies from different skin cancer types were analyzed for the study of cutaneous HPV types, alone or in parallel, with samples from healthy skin as a comparison. The researcheuthors have used only molecular biology techniques, beginning with classical PCR, followed by hybridization, qPCR, multiplex genotyping, and NGS. The primers used were targeting the E1 β-HPV gene fragment, two pairs of general degenerate primers CP65–70 (CP65/70 and CP66/69, consensus primer pair FAP (FAP59\FAP64) targeting the 5′end of the L1 ORF, FAP and PGMY-GP + primer systems, and E7 gene for HPV types. The analyzed studies found HPV types in different percentages in skin cancer and newly identified HPV types were reported. The researcheauthors underline the need of optimizing the sensitivities of the used assays and the necessity of confirmatory methods ^{[41][42][43][44][45]}[41,42,43,44,45].​​​​​​​